Isolation and Characterization of Novel Strains of <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> and <Emphasis Type="Italic">Serratia marcescens</Emphasis> Possessing High Efficiency to Degrade Gasoline,Kerosene, Diesel Oil,and Lubricating Oil

Bacteria possessing high capacity to degrade gasoline, kerosene, diesel oil, and lubricating oil were screened from several areas of Hokkaido, Japan. Among isolates, two strains, WatG and HokM, which were identified as new strains of Pseudomonas aeruginosa and Serratia marcescens species, respectively, showed relatively high capacity and wide spectrum to degrade the hydrocarbons in gasoline, kerosene, diesel, and lubricating oil. About 90-95% of excess amount of total diesel oil and kerosene added to mineral salts media as a sole carbon source could be degraded by WatG within 2 and 3 weeks, respectively. The same amount of lubricating oil was 60% degraded within 2 weeks. Strain HokM was more capable than WatG in degrading aromatic compounds in gasoline. This strain could also degrade kerosene, diesel, and lubricating oil with a capacity of 50-60%. Thus, these two isolates have potential to be useful for bioremediation of sites highly contaminated with petroleum hydrocarbons.     

Loss‐of‐function of ASPARTIC PEPTIDASE NODULE‐INDUCED 1 (APN1) in Lotus japonicus restricts efficient nitrogen‐fixing symbiosis with specific Mesorhizobium loti strains

The nitrogen‐fixing symbiosis of legumes and Rhizobium bacteria is established by complex interactions between the two symbiotic partners. Legume Fix^– mutants form apparently normal nodules with endosymbiotic rhizobia but fail to induce rhizobial nitrogen fixation. These mutants are useful for identifying the legume genes involved in the interactions essential for symbiotic nitrogen fixation. We describe here a Fix^– mutant of Lotus japonicus, apn1, which showed a very specific symbiotic phenotype. It formed ineffective nodules when inoculated with the Mesorhizobium loti strain TONO. In these nodules, infected cells disintegrated and successively became necrotic, indicating premature senescence typical of Fix^– mutants. However, it formed effective nodules when inoculated with the M. loti strain MAFF303099. Among nine different M. loti strains tested, four formed ineffective nodules and five formed effective nodules on apn1 roots. The identified causal gene, ASPARTIC PEPTIDASE NODULE‐INDUCED 1 (LjAPN1), encodes a nepenthesin‐type aspartic peptidase. The well characterized Arabidopsis aspartic peptidase CDR1 could complement the strain‐specific Fix^– phenotype of apn1. LjAPN1 is a typical late nodulin; its gene expression was exclusively induced during nodule development. LjAPN1 was most abundantly expressed in the infected cells in the nodules. Our findings indicate that LjAPN1 is required for the development and persistence of functional (nitrogen‐fixing) symbiosis in a rhizobial strain‐dependent manner, and thus determines compatibility between M. loti and L. japonicus at the level of nitrogen fixation.     

Fruiting phenology of animal-dispersed plants in response to winter migration of frugivores in a warm temperate forest on Yakushima Island, Japan

The fruiting phenology of animal-dispersed plants was observed in a warm temperate, evergreen forest on Yakushima Island. The number of ripe fruits was counted for 22 trees, four lianas and one parasitic epiphyte species with sapfruit. These fruits were consumed by birds and Japanese macaques (Macaca fuscata yakui). Birds with small gapes (e.g. Japanese white-eye [Zosterops japonica]) consumed only small fruit less than 6 mm in diameter, while birds with large gapes (e.g. red-capped green pigeon [Sphenurus formosae]) and Japanese macaques consumed a wide range of fruits from 4 to 16 mm in diameter. The larger animals did not ignore the smaller fruits. Brown-eared bulbul (Hypsipetes amaurotis) and Japanese white-eye were the main consumers of sapfruit in terms of frequency in winter. Some of the observed consumers were year-round residents, but most of the consumers migrated to Yakushima Island from the main islands of Japan to overwinter (from November to March), and their abundance in winter was four times as high as during the rest of the year (from May to October). In 23 of the 27 plant species investigated, sapfruit production coincided with their immigration season, whereas tree species bear capsules and nuts during autumn from September to November. We suggest that sapfruit species set their ripe period to the season when frugivorous birds are most abundant.     

15-Cis-β-carotene found in the reaction center of spinach Photosystem I

-Carotene was extracted from spinach Photosystem I reaction centers (one consisting of the Psa A, B, C, D and E subunits and the other consisting of the Psa A and B subunits alone), and the extract was subjected to high-pressure liquid chromatography using an apparatus equipped with a two-dimensional diode-array detector; all the procedures were performed at 4 °C in complete darkness. Both 15-cis and all-trans--carotene were identified in the extract by means of electronic absorption spectroscopy. Thus, universal presence of 15-cis carotenoid in the reaction centers of purple photosynthetic bacteria and of spinach Photosystem I and Photosystem II has been shown.Abbreviations Chl- chlorophyll - DEAE- diethylaminoethyl - DMF- dimethylformamide - HPLC- high pressure liquid chromatography - LHC- light-harvesting complex - PS- Photosystem - RC- reaction center - RC_a,b- reaction center consisting of Psa A and B subunits alone - RC_a-e- reaction center consisting of Psa A, B, C, D and E subunits     

Effect of second-messenger modulators in K-562 cell differentiation: dual action of calcium/phospholipid-dependent protein kinase in the process of differentiation

We investigated the roles of second messengers in K-562 cell differentiation induced by either commitment-inducing agents (Ara-C, thymidine), or a noncommitment-inducing agent (hemin). Cell differentiation induced by both types of agents was inhibited by dbc-AMP, staurosporine, and H-7. In contrast, OAG enhanced hemin-induced cell differentiation, but it inhibited that due to Ara-C or thymidine. When K-562 cells were incubated with 4 x 10(-6)M hemin or 2 x 10(-7)M Ara-C for 2 days, an increase of epsilon-mRNA occurred. The addition of cycloheximide (1 microgram/ml) completely blocked this change, suggesting that de novo protein synthesis was necessary for the increase of epsilon-mRNA. Simultaneous treatment with Ara-C and cycloheximide for 2 days did not block either the increase of epsilon-mRNA or that of benzidine-positive cells, which were measured after 5 days of further incubation without additives. This suggested that the process of Ara-C-induced K-562 cell differentiation could be divided into two steps, i.e., a commitment step and a phenotypic expression step, and that the commitment step was at least partly resistant to cycloheximide. We investigated the roles of second messengers in each step. Our results suggested that PKC may act as a negative regulator of commitment step and as a positive regulator of the phenotypic expression. This may explain the differing effects of OAG on hemin- and Ara-C-induced K-562 cell differentiation.     

Preparation of active hybrid enzymes composed of the native and chemically inactivated aspartase subunits from Escherichia coli

The hybridization of the native and chemically inactivated aspartase from Escherichia coli was studied. Preparations of the tetrameric enzyme obtained by mixing the native and N-ethylmaleimide (NEM)-inactivated aspartase in 4 M guanidine-HCl followed by 51-fold dilution at room temperature retained catalytic activity. Affinity chromatography on AF-Red TOYO-PEARL separated several active components in the hybridized preparations, and the presence of [14C]NEM-inactivated subunits in the active hybrids was demonstrated. The addition of the native aspartase to Sepharose-bound NEM-inactivated enzyme in 4 M guanidine-HCl resulted in the formation of an immobilized enzyme with enzyme activity. The specific activity of the various hybrids, composed of unmodified and [14C]NEM-inactivated subunits, was roughly proportional to the number of unmodified subunits in each tetramer. Furthermore, when reversible denaturation was conducted on mixtures of the native and NEM-inactivated enzyme at various proportions, the enzyme activity recovered was proportional to the amount of the native enzyme added. These results strongly suggest that each subunit makes an independent contribution to the overall enzyme activity regardless of the presence of other subunits in the same molecule. The theoretical and practical implications of this work are discussed.     

Characterization of multiple fumarase proteins in Escherichia coli

Two different types of fumarase were found in sonic extracts of Escherichia coli; one required Fe-S for the enzyme activity, and the other did not. When the cells were grown without aeration, the Fe-S-independent enzyme occupied over 80% of the overall fumarase activity. Highly purified Fe-S-independent enzyme was suggested to be composed of four subunits (Mr = 48 kDa) by SDS-polyacrylamide gel electrophoresis and gel filtration. Amino acid and N-terminal sequence analyses supported the possibility that the enzyme is a product of fumC gene (FUMC). In aerobically grown cells, however, the content of FUMC was low and the Fe-S-dependent fumarase occupied over 80% of the overall activity. The Fe-S-dependent enzyme appeared to be labile and the activity was rapidly lost during purification. Although the spontaneous inactivation was previously ascribed to thermal lability (S.A. Woods & J.R. Guest (1987) FEMS Microbiol. Lett. 48, 219), the activity could be restored by anaerobic incubation with ferrous ions and SH-compounds.     

Evidence of tool-use by chimpanzees (Pan troglodytes schweinfurthii) for digging out a bee-nest in the Kahuzi-biega national park,Zaïre

Two sticks were found near a broken bee-nest ofMeliplebeia tanganyikae aff.nigrita Alfken in the Mt. Kahuzi region of Zaïre, and were thought to have been used by a chimpanzee or perhaps several chimpanzees to dig out the subterranean nest. Honey, larvae, and most of the nest had been eaten by them. We did not find any evidence to indicate tool-use by chimpanzees in the Masisi or Itebero-Utu regions, although stingless bees were observed and honey was eaten by chimpanzees in both regions. The sticks resembled in length and diameter those known to be employed for digging termite-mounds in south-west Cameroon and Equatorial Guinea. The tool-behavior of the chimpanzees observed at Mt. Kahuzi may be similar to that of those in central Africa, rather than of those in east Africa where digging-tools have yet to be found. Another possibility is that the chimpanzees have developed the digging-tools independently, based on the need to take animal protein in the Mt. Kahuzi region, where termite-mounds are rarely observed. Instead of seeking termites, they may have a stronger motivation to seek bee larvae, especially the larvae of stingless bees beneath the ground, than to the chimpanzees inhabiting lower or drier forests.